Okumura H, Arashima M, Ohuchi J, Kasai Y, Tsukumo K, Kakishima H, Kotani M, Kojima H, Kurishita A, Hayashi M, Miyajima A, Sunouchi M, Ohno Y
Japan Cosmetic Industry Association, Hatsumei Bldg, 9-14, Toranomon 2-chome, Minato-ku, Tokyo, 105-0001Japan; NOEVIR Co., Ltd, 112-1 Okada-cho, Yokaichi, Shiga, 527-8588Japan.
Toxicol In Vitro. 1999 Feb;13(1):199-208. doi: 10.1016/s0887-2333(98)00074-5.
The present interlaboratory validation study was performed in order to evaluate the use of Chinese hamster lung cell lines that employs crystal violet staining (CHL-CVS) as an alternative cytotoxicity test to the Draize eye irritation test (Draize test) for cosmetic ingredients. Ten substances, nine of which were surfactants, were evaluated at seven laboratories in the first phase of the validation study; 15 substances including dyes and lipids were evaluated at seven laboratories in the second phase of the validation study; 14 substances including acids and alkalis were evaluated at four laboratories in the third phase of the validation study. The logEC(50) values obtained for CHL-CVS were compared with the maximal average Draize total score (MAS) for a 10% (w/v) solution of 38 cosmetic ingredients as well as isotonic sodium chloride solution. The interlaboratory coefficient of variation (CV) for EC(50)s was 35.6%, which was considered to be within a tolerable range. The correlation coefficient and the Spearman's rank correlation coefficient between the in vitro and in vivo tests were -0.729 and 0.709, respectively. The prediction ability of the proposed method was assessed from the linear regression line for a MAS cut-off point of 15. According to this analysis, four substances (two alcohols and two acids) were determined to be false negative. The present study revealed the following characteristic factors of this method: (1) CHL-CVS could be applied to all the test substances including dyes and lipids in this study; (2) The results for medium-insoluble substances varied according to the laboratory; (3) The correlation between the in vivo and in vitro data for acids and alcohols (lower mono-ol) differed from that of the other substances. These results suggested that the CHL-CVS might have a potential to predict the Draize MAS if definite criteria can be established for the compounds to be applicable.
进行本次实验室间验证研究的目的是评估采用结晶紫染色的中国仓鼠肺细胞系（CHL-CVS）作为化妆品成分的替代细胞毒性试验，以替代兔眼刺激试验（Draize试验）。在验证研究的第一阶段，七个实验室对十种物质进行了评估，其中九种为表面活性剂；在验证研究的第二阶段，七个实验室对包括染料和脂质在内的十五种物质进行了评估；在验证研究的第三阶段，四个实验室对包括酸和碱在内的十四种物质进行了评估。将CHL-CVS获得的logEC(50)值与38种化妆品成分的10%（w/v）溶液以及等渗氯化钠溶液的最大平均Draize总分（MAS）进行比较。EC(50)的实验室间变异系数（CV）为35.6%，被认为在可接受范围内。体外和体内试验之间的相关系数和Spearman等级相关系数分别为-0.729和0.709。根据MAS截止点为15的线性回归线评估了所提出方法的预测能力。根据该分析，确定有四种物质（两种醇和两种酸）为假阴性。本研究揭示了该方法的以下特征因素：（1）CHL-CVS可应用于本研究中的所有受试物质，包括染料和脂质；（2）中等不溶性物质的结果因实验室而异；（3）酸和醇（低级一元醇）的体内和体外数据之间的相关性与其他物质不同。这些结果表明，如果能够为适用的化合物建立明确的标准，CHL-CVS可能有预测Draize MAS的潜力。
